Civilians in the US and other countries face a growing threat from bioterrorism. It is not feasible to develop effective vaccines or treatments for all possible biowarfare agents that may be encountered. Furthermore, the identity of the bioterror agent(s) employed in an attack may not be known in time to administer specific therapies. Therefore, it is highly desirable to develop broad-spectrum nonspecific therapies that can protect against a wide range of pathogens. In theory, pre-activating the appropriate innate defenses should reduce morbidity and mortality, and improve the response to supportive therapy. Hundreds of millions of years of evolution have endowed humans with an extraordinary range of innate immune defenses against viruses, bacteria, fungi, and parasites. This innate immunity is thought to depend in large part on a family of molecules called "Toll like receptors" (TLRs), which bind molecules that are present in microbes, but not in normal host tissues. Recent studies show that ligands for TLR9, called CpG motifs, are particularly effective at activating broad-spectrum innate immune defenses, and can protect animals against a wide range of pathogens. Preclinical studies show that pretreatment with a synthetic CpG oligodeoxynucleotide (ODN) protects mice against a wide variety of viruses, bacteria, and intracellular parasites, including the Category A agents anthrax, tularemia, and Ebola. For protection against an inhaled pathogen, the optimal route of administration for the CpG ODN also appears to be inhaled. Three classes of CpG ODN have been identified, CpG-A, CpG-B, and CpG-C, with distinct immune profiles with human cells in vitro and in vivo in mice. Phase I/II human clinical trials in over 300 subjects administered a CpG-B ODN suggest strong enhancement of immune responses and an excellent safety profile. The overall goal of this proposal is to identify a CpG ODN that provides broad-spectrum defense against pathogens in animal challenge models, and to develop this CpG for protection of civilians against a bioterror agent. Based on studies of over 3000 ODN in the three classes, one optimized ODN from each class has been selected for comparison in this program: CpG-A 10108, CpG-B 10105, and CpG-C 10107. Our proposal is based on the hypothesis that the prophylactic activation of particular innate immune defenses with a CpG ODN will protect against a subsequent exposure to one or more bioterror agents. It is expected that pretreatment with CpG would reduce morbidity and mortality from bioterror and prolong the "golden hour" during which supportive therapy and specific countermeasures could be administered. If successful, the proposed studies would provide supportive data for human safety clinical trials and an eventual NDA submission. This goal will be accomplished through the following specific aims: Specific Aim 1. Determine the efficacy of different CpG classes in protecting mice against inhaled Category A pathogens. Specific Aim 2. Determine the efficacy of different CpG classes in protecting against other inhaled pathogens. Specific Aim 3. Determine the effect of inhaled CpG classes on respiratory tract immune function. Specific Aim 4. Identification of peripheral biomarkers for inhaled CpG ODN (mice, swine, sheep, primates). Specific Aim 5. Characterize the toxicity of inhaled CpG ODN. Specific Aim 6. Characterize the absorption, distribution, metabolism, and excretion of inhaled CpG ODN.